CA2018067A1 - Acylated derivatives of etoposide - Google Patents
Acylated derivatives of etoposideInfo
- Publication number
- CA2018067A1 CA2018067A1 CA002018067A CA2018067A CA2018067A1 CA 2018067 A1 CA2018067 A1 CA 2018067A1 CA 002018067 A CA002018067 A CA 002018067A CA 2018067 A CA2018067 A CA 2018067A CA 2018067 A1 CA2018067 A1 CA 2018067A1
- Authority
- CA
- Canada
- Prior art keywords
- compound
- etoposide
- group
- benzoyl
- alkanoyl
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Abandoned
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Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
- C07H15/00—Compounds containing hydrocarbon or substituted hydrocarbon radicals directly attached to hetero atoms of saccharide radicals
- C07H15/20—Carbocyclic rings
- C07H15/24—Condensed ring systems having three or more rings
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
- C07H17/00—Compounds containing heterocyclic radicals directly attached to hetero atoms of saccharide radicals
- C07H17/04—Heterocyclic radicals containing only oxygen as ring hetero atoms
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P35/00—Antineoplastic agents
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- General Health & Medical Sciences (AREA)
- Engineering & Computer Science (AREA)
- Biochemistry (AREA)
- Biotechnology (AREA)
- Genetics & Genomics (AREA)
- Molecular Biology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Medicinal Chemistry (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Pharmacology & Pharmacy (AREA)
- Animal Behavior & Ethology (AREA)
- Public Health (AREA)
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- Saccharide Compounds (AREA)
- Heterocyclic Carbon Compounds Containing A Hetero Ring Having Oxygen Or Sulfur (AREA)
Abstract
ABSTRACT
The epipodophyllotoxin glucosides disclosed are acylated at herein one or both sugar hydroxyl groups.
These compounds exhibit significant activity against P388 murine leukemia.
The epipodophyllotoxin glucosides disclosed are acylated at herein one or both sugar hydroxyl groups.
These compounds exhibit significant activity against P388 murine leukemia.
Description
Z~ 67 B~CKGROUND O~ THE INVENTION
1. Field of the Invention The present invention relat.~s to derivati;es of epipodophyllotoxin, their use as antitumor agen's, and pharmaceutical compositions containing them. More particularly, the no~el compo~ ds of the present inve!ltion are acyl derivatives of 4'-demetl~ylepipodopllyllotoY.i glucosides.
1. Field of the Invention The present invention relat.~s to derivati;es of epipodophyllotoxin, their use as antitumor agen's, and pharmaceutical compositions containing them. More particularly, the no~el compo~ ds of the present inve!ltion are acyl derivatives of 4'-demetl~ylepipodopllyllotoY.i glucosides.
2. Description cf Background Art Etoposide and teniposide are clinically useful anti~tumor agents derived from the naturally occurring lignan, podophyllotoxin. Currently etoposide is marketecl in the United States under the trade name Vepesid for the treatment of small cell lung cancer an~ testicular cancer.
The favorable pharmacological properties of etoposide and teniposide have encouraged much activity in the search fo-other active analogs within the same class.
4'-0-Demeti~ylepiF~dophyllotoxin glucosides whereill the hydroxyl gro~lp~ of the ~ugar moiety are acylated ha~e been reported ir. the literat~lre as intermediatPs for the preparation of tlle correspondincs 4'-demetllylepipodophyllo-toxin glucosides; howe~-er, the phell5l a;OUD of ~hese compounds are also protected.
Canadian Patent No. 956,939 cliscloses compounds of formula (I~
? ZS~18067 R l ~o~\ O
<~ o H3C~OCH3 o-C-ocH2-R3 ( I ) wherein R1 is C1 5 alkyl; R2 is acetyl or formyl; and ~3 is phenyl or substituted phenyl; possible substituted phenyls mention~d but not exemplified are p-nitrophenyl and p-meth~.~xyphenyl.
U.S. Patent 4,564,675 discloses compounds of the formula (II) H 3 C--~ o R
<~Q
H3C~OCH
O-- R
I I ) ~herein R is -C(O)CH~X, X is a haloge.. ztom.
European Patent Appli^~tion 162,701 discloses compounds of formula (III) Z~8g:~7 <ô 1~ ~0 H3CO~OCH3 RZ
(111) wherein Rl and R2 may be the same or different and each represent -C(O)CHX2 or -C(O)CX3 wherein X is a halogen atom.
Japanese Kokai 58/225,096 (Derwent Abst. No.
84-034268/06) and 58/219,196 (Derwent Abst. No.
84-027495/05) disclose compounds of form~lla (IV) and (V), respectively.
CH3~ " CH3--~,~0, o ~0 ~ ~'' 1 ~ X < ~
H3CO~OCH3 H3CC$oCH3 P
( lV) (~') wherein A stands for -C02-CH2-C(H)m(X)n wherein X is a halog~n atom and m is 0 to 2 and n is 1 to ~, m+n=3, and Ac is acyl.
2~ 6~
European Patent hpplication 226,202 discloses an intermediate for etoposide synthesis having the formula (VI) O' ,~
<~
H3C(~OCH3 R
~ V I ) wherein A represents an acetyl group.
Mono-hemisuccinate derivatives of etoposide having the formuias (VII) are reported in J. Pharm. Biomed, ~nal., 1987, 5(1)~ 20 3--t;;~,~o `~6 ~3c~ocl13 (vll) wherein one of R1 and R2 is H and the other is -CO(CH2)2C02H. These compounds are used as a means to conjugate etoposide to bovine serum albumin.
- ! zn~
The favorable pharmacological properties of etoposide and teniposide have encouraged much activity in the search fo-other active analogs within the same class.
4'-0-Demeti~ylepiF~dophyllotoxin glucosides whereill the hydroxyl gro~lp~ of the ~ugar moiety are acylated ha~e been reported ir. the literat~lre as intermediatPs for the preparation of tlle correspondincs 4'-demetllylepipodophyllo-toxin glucosides; howe~-er, the phell5l a;OUD of ~hese compounds are also protected.
Canadian Patent No. 956,939 cliscloses compounds of formula (I~
? ZS~18067 R l ~o~\ O
<~ o H3C~OCH3 o-C-ocH2-R3 ( I ) wherein R1 is C1 5 alkyl; R2 is acetyl or formyl; and ~3 is phenyl or substituted phenyl; possible substituted phenyls mention~d but not exemplified are p-nitrophenyl and p-meth~.~xyphenyl.
U.S. Patent 4,564,675 discloses compounds of the formula (II) H 3 C--~ o R
<~Q
H3C~OCH
O-- R
I I ) ~herein R is -C(O)CH~X, X is a haloge.. ztom.
European Patent Appli^~tion 162,701 discloses compounds of formula (III) Z~8g:~7 <ô 1~ ~0 H3CO~OCH3 RZ
(111) wherein Rl and R2 may be the same or different and each represent -C(O)CHX2 or -C(O)CX3 wherein X is a halogen atom.
Japanese Kokai 58/225,096 (Derwent Abst. No.
84-034268/06) and 58/219,196 (Derwent Abst. No.
84-027495/05) disclose compounds of form~lla (IV) and (V), respectively.
CH3~ " CH3--~,~0, o ~0 ~ ~'' 1 ~ X < ~
H3CO~OCH3 H3CC$oCH3 P
( lV) (~') wherein A stands for -C02-CH2-C(H)m(X)n wherein X is a halog~n atom and m is 0 to 2 and n is 1 to ~, m+n=3, and Ac is acyl.
2~ 6~
European Patent hpplication 226,202 discloses an intermediate for etoposide synthesis having the formula (VI) O' ,~
<~
H3C(~OCH3 R
~ V I ) wherein A represents an acetyl group.
Mono-hemisuccinate derivatives of etoposide having the formuias (VII) are reported in J. Pharm. Biomed, ~nal., 1987, 5(1)~ 20 3--t;;~,~o `~6 ~3c~ocl13 (vll) wherein one of R1 and R2 is H and the other is -CO(CH2)2C02H. These compounds are used as a means to conjugate etoposide to bovine serum albumin.
- ! zn~
4'-Phosphate of etoposide and the disodium salt thereof are disclosed in Japanese Kokai 63/192,793.
CH~ ~
~ H ~..
~0 <~ ~"'\~
CU3~0CU3 ~ ~oeos!de_ehoseha ~e SUMM~R~' OF TEE INV~NTION
The present invention provides compounds having the formula (VIII) R L~
(~ ~0 CH~ CH3 ORS
~Vl I I ) wherein R2 is H and Rl is selected fro~ the group consisting f (Cl-lo)alkyl~ (C2-lO)alkenylr ~C5 6)cycloalkyl~ 2-furyl, y , ( 6_l0)aryl, and (C7_14)aralkyl; or R and ~ a~-e each (Cl lO)alkyl; or Rl, R and the carhon to which they are at_ached together represent ~C5 6)cyclo21kyl; one of R
and R^ is ~ and the other is selected from the group 2~ 6~ ~
consisting of (Cl 5)alkanoyl and benzoyl; or R3 and R4 are the same and are selected from the group consisting of (C1 5) alkanoyl and benzoyl; R5 is H or a phosphate group.
Another aspect of the present invention provides pharmaceutical compositions comprising an antitumor effective amount of a compound of formula (VIII) and a pharmaceutically acceptable carrier.
Yet a further aspect of the present invention provides a method for inhibiting tumor growth in a tumor bearing mammalian host which comprises administering to said hcst an antitumor amount of a compound of form~lla (VIII).
DET~ILED DESCr~IPTION OF `1~ INVENTION
A preferred embodiment of compounds of formula (VIII) comprises those compounds wherein R2 is H and ~1 is selected from the group consisting of methyl, 2-thienyl, and phenyl, with methyl being the most pleferred substituent.
A further preferred embodiment pro~i.des _ompounds of formula (VIII) wherein R3 and R4 are each ~C1 5~alkanoyl; a most preferred embodiment provides compounds of formula (VIII) wherein R3 and R4 are each a formyl gro~lp.
Yet a further preferred embodiment pro~,ides compounds of formula (VIII) where-in one of h- and R4 is H, and the other is (Cl 5~alkanoyl or benzoyl.
As used herein, the term "phosphate".ir.~ludes the-group-PO~H2 and pharmaceutically acceptable salts tilereof.
Z()1~7 -~
Pharmaceutica'1y acceptable salts include both monobasic and dibasic salts wherein the cation includes, but not limited to, alkali metals, e.g. sodium, potassium and lithium;
alkaline earth metals, e.g. magnesium, calcium and barium;
orgnic amine salts, e.g. ammonium.
The novel compounds of the present invention may be prepared by reactin~ a 4'-phenol protected 4'-demethylepipodophyllotoxin g]ucoside o formula ( X~) with the desired carboxylic acid or an acylating equivalent thereof, followed by removal of the phenol protecting group.
R ~1~\ D
<~0 CH3 ~OCH3 y ~ a: Y ~ o ~ l i ng gl- oup lXb: Y = H
~ he phenol protecting group is not par~icularly restricted and may be any that can be irtroduced and removed with little or no adverse effect on other parts of the molecule. More particularly, the phenol protecting g~oup should be removable by methods that ~c not distu-b the acyloxy functionalities cn the sugar moiety o- the final produc~s. Methods fcr phenol protection a~-e well kno~r. in the art and as examples of aeneral classes of phenol protecting groups, mention can be made o~ ethers, acetzls, esters, and carbonatec. For the purpose of the present invention, we found the bPn~yloxycarbonyl racliczl mzy be 6~ ~, conveni~r_ly so employed. Thus by reacting a 4'-demethyl-epipodophyllotoxin glucoside of formula (IXb) with benzylchloroformate at low temperature, e.g. -15C, in an inert organic solvent such as methylene chloride and in the presence of a tertiary organic base such as pyridine, the corresponding 4'-0-benzylo~ycarbonyl-4'-demethylepipodo-phyllotoxin glucoside is obtained. Compounds of formula (IXb) and their synthesis are disclosed in US Patent No.
3,524,844.
The acylation of the sugar llydro~yl groups of compounds of formula (IXa) may be effected using the desired carboxylic acid or an acylating agent derived therefrom, for example an acid halide such as acid chloride, active ester derived from N-hydroxysuccinimide or 1-hydroxytriazole, and symmetrical or mixed anhydride. When the carboxylic acid is used as the acylating species, it is preferably used in conjunction with a condensing agent for e~ample a carbodiimide such as dicyclohexylcarbodiimide (DCC).
Acylating 2gents are C1 5alkanoic acids and ber.~oic acid and acylating equivalents thereof as above d~scribed. Alkanoic acids include, but are not limited to, formic acid, acetlc acid, propionic acid, butyrlc acid, pentanoic acid, and branched alkanoic acids such as 2-methylpropionic acid and 3-methylb~_tanoic acid.
The acylation reaction may be carried out in an inert organic solvent such as pyridine, and prefera~ly in-lude n the reaction mixture an acid acce?~o~ ~hen acid is ex?ec~ed to be a by-?roduct; suitable acia acceptors are for example tertia-y amine bases such as pyri-ine, triethylamine, diisopropylethylamine and the like, or inorganic bases such as sodium anG potassium carbonates. The reaction may be conGucted a~ tempera~ures conduci~-e to forma_ion of the Z5)~6~ -~
~sired products an~ may be from about -15C to about 50C;
the reaction may ta~e several minutes to several days foi-completion deppending on the nature of the reactants and choice of reaction conditions such as temperature.
The acylation is not regioselective and may result in a mixture of 2"-monoacylated product, 3"-monoacylated product and 2",3"-bisacylated product. However, when at least two equivalents of the acyla,ing agent is used relative to the epipodophyllotoxin reactant, the bis-acylated derivative is produced predominantly. The proportion of monoacylated product and bisacylated product may be controlled to some extent by manipulative reaction parameters according to principles commonly practiced in the art, e.g. varying the relative amounts of reactants, or varying reaction conditions such as temperature and length of reaction time.
In general when equimolar of the reactants are e~..loyed, reduced reaction temperature and shorter reaction time tend to increase the amount of monoacylated products formec.
Alternatively, monoacylated products may be obtained by first protecting one of the sugar hydroxyl groups of the 4'-phenol protected epipodophylloto~in glucoside starti;.g material. The protecting group for the sugar hydroxyl group is preferably a bul~y protecting group, for example the t-butyldimethylsilyl group, in order to minimize derivatizing both hydroxyl groups. Again, protection of the sugar hydroxy-l groups is not regioselective; however, if desi-ed, the 2"-protec'ed cr 3"-prot~_ted compounds are easily separa~ed chroma.og aphically. We have carried ou formylation reactions of eto?oside; howe~er, we have no~
been slccessful in isola~ing either of ~he expected mono ormyl derivatives of etoposide.
Z01~3~6~7 The product mixture comprising pl~enol-protected acylated derivatives may be separated into individual components using conventional separat;on techniques such as column chromatography. Deprotection of the phenolic hydroxyl group and, if necessary, the sugar hydroxyl gro-lp provides compounds of the present invention. The deprotection may be effected prior to or subsequent to the isolation of the individual components; the order is not critical. The protecting group i~ removed by usin~ methods suitable for the protecting group selected and these are generally well known in the art. For example, when the protecting group is benzyloxycarbonyl, it may be removed by catalytic hydrogenation at atmospheric precsure using palladium on carbon as the catalyst; the t-butyldimethyl-silyl group may be removed by heating in an aqueous alcoholic solution or by treating with fluoro anion or aqueous acetic acid.
Following the removal of the phencl protecting group, the ~'-phenol g~oup may be further derivati~ed to provide the 4'-phosphate of formula (VIIl). Phosphorylation of the 4'-phenol may be carried out using conventional methods.
For example, a compound of formula (VIII) wherein R5 is H
may be reacted with a phosphorylating agent such as phosphorous o~ychloride; the phosphoryl chloride intermediate i~ hydrolyzed i_ situ to give the 4'-pllosphate.
Hydrolysis in the presence of a base e.g. sodiu~ bicarbonate provides the corresponding salt. This procedure as well as other potentially applicable phosphorylation methods is disclosed in U.S. patent application GB2,207,674 which is hereby incorporated by refe ence.
201~3~6'7 It is to be understood that synthesis of compounds of the present invention is not limited to the procedures and reagents outlined above, but may include other methods capa~le of acylating the hydroxyl groups on the sugar portion of 4'-demethylepipodophyllotoxin glucosides. The reaction conditions will of course vary with the choice of startlng materials but may be ascertained by a skilled artisan without und~le experimentation.
BIOLOGICAL ACTIVITY
Representat-ive compounds of the present invention were evaluated for antitumor activity against murine transplantable P38~ le~kemia. Female CDF1 mice were inoculated intraperitoneally with 0.4 ml of diluted ascitic fluid containing 106 lymphocytic leukemia P388 cells. Test compounds were administered intraperitoneally as a .single dose on day 1 and animals were observed for 50 days. The percent increase of median survival time (MST) of treated animals over that of un~reated control animals was determined and re?orted as ~O T/C. Compounds sho-;ing % T/C
values of 125 or greater are considered to have significant antitumor activity. Table I presents the results of the in vivo evaluation; only th~ maximum ,O T~C and the dose giving the maximum effects are reported.
~O~ 7 Table I. Antitumor activitv _f~a1nst P3~8 leukemia Compound _o e_~mg~kgf/day~ ~T/C MST
Example 1 30 165 Example 3 30 240 Ex~mple 4 30 210 Etoposide 30 182 Example 6 120 >500 ~3/4)*
Example 8 120 14r`
Example 9 60 f~20 ( l/f~ ) Example 10 120 125 Etoposide 120 >390 (6/12) *Number of survivors/tested on day S0 ~/~ /S/
In vitro cytotoxicity of compounds of the presen-t R G invention was evaluated against ' ]~r c~ ' l r~ r ~f~ B16-F10 murine melanoma ~ oser human colcn carcinoma~
bl lT '~ ~ ~;r r ~ r r r ~ ~ ~ ~ I~r~ ~ ^ T~
r^r- ~t~ -1 i nf~ ~ - r~ r ~~ L - r~
Iql; E R/6~ o f~L Mo~J
n~(f -~ I~C~ ~f ~xponentially gr ~ e harvested, counted and suspended in the culture me~dium at the ~; concentrations of 1.5 x 104 and 3 x 104 cells/ml, respectively. Twenty-four hours after p].anting c~11 suspensicn (î80 mcl) into wells of a rJo-weil microti~er plate, test materia s (20 mcl) were added to the wells and the ?lates were incubated for /2 hours. The cyto oxi_ activi~ies against ~he tum~r cells were colorimet~-ically 2(~ 7 determined at 540 nm after staining viable cells with ~/~ ~&~ neutral red solution. ~or cytotol~ic-it~ pcrimcn~-~
R ~ K56. ~nd ~56~ DM ccllc, ~00 m~l-ef thc ccll ~u3~cnsi~
~,~k,~ 104 ccll-~ml) ~ac incub~ted ~.~ith tcct m~e~al3 ~100 mel) a' 37C, 5,~ C~2 for 4~ hollr~ in - 4 ~cll tis~uc cult~e-p~
~hc c~toto~ic actir'~ Jas-~e~crmincd b~ e~unt~ e num~cr of colls uEing a cell ~ ntcr (s~Gme~ cc 120.~). The results are summarized in Table II.
~/z6/~/ u~Y~ ~ n~ ~JQ~
Table II. In vitro cytotoxicity_~a_nst ~ariouL ce'~ lin~
h~
~l');
IC50 (mcg~
Compound B16-F10 Moser~ I~rC ,'~
Example 3 1.6 1.1 0.
~/6/~ Example 4 1.4 ND* ~D ~
Example l 1.3 <0.8-0.1~ ~ ' `50 Example 8 0.92 ND~D ,~\ ~ID
Example 9 0.17 0.83O~4~J/~ \\ 6.7 Example lO 4.4 ND I~
Example 6 2.0 0.84 Etoposide 0.45 ND 'O;O
ND: Not determined Accordingly, the present invention provides a method ror inhibiting mammalian tumors ~hich comprises aaministering an effective tumor-inhibiting dose of an antitumor co~pound of formula VIII to a tumor bearing host.
For this purpose, the druc may be admin,stered by conventional routes including, but not limited to, intravenous, intramuscular, intratumoral, intraarterial, ~8~67 ^ ~
intralymphc~ic, and oral.
A further aspect of the present invention provides a pharm2-eutical composition whi.ch comprises a compound of formula VIII and a pharmaceutically acceptable carrier. The antitumor composition may be made up of any pharmac~-~ti-cal form appropriate for the desired route of administra-tion. Examples of such compositions include solid compositions fGr oral administration such as tablets, capsules, pills, powders and granules, liquid compositlons for oral administration such as solutions, suspensions, syrups or elixirs and preparations for parenteral administration such as sterile solutions, suspensions or emulsions. They may also be manufactured in the form of sterile solid compositions which can be dissolved in sterile water, physiological saline or some other sterile injectable medium immediately before use.
Optimal dosages and regimens for a givPn mammalian host can be readily ascertained by those skilled n the art. It will, of co~rse, be ap~reciated that the actual dose used will vary according to the particu].ar co.m?osition formulated, the partic~lar compound used, the mode of application and the particular site, host and disease being treated. Many factors that modify the action of the drug will be taken into account incl~lding age, weight, sex, diet, time of adm.inistration, rou~e of administra~ion, rate o.
e~cretion, condition of the patien', drug combinatiolls, reaction sensiti~;ities and sP~erity of the disease.
The following exam?les a-e for illus~rative pu-?oses only and should not be construPd as limiting the _cope of the invention which is defined solely by the Claims appended to the specification.
,;~ a67 Preparation of 4'-O-Benzyloxycarbonyl Etoposide (4'-CBZ
etoposide) Benz~ lchloroformate (1.98 ml, 15 mmol) was addeP over a period of 30 minutes to a mixture of etoposide (5.88 g, 10 mmol) and p~,ridine (10 ml) in methylene chloride (100 ml) kepl; at -15C. The mixture was stirred at -15C for an additional ho~lr, washed successively ~ith 5% HCl, aqueous sodium hicarbonate, and water, and then driec o~er anhydrGus sodium sulfate. The solvent was evapora1:ed to gi~e 8.05 ~3 of the crude product which ~,~as purified on a silica gel column (5% methanofl-methylene chloride) to give 6.93 g (96%) of 4'-O-benzyloxycarbonyl etoposide as a colorless semi-solid. M.P. 152-155C
IR ~max (Nujol) cm : 320"0-3600 (OH), 1760 (lactone &
4'-0-benzyloxy carbonyl), 1600 (aromatic).
H NMR (60 MHz, CDCl_) ~ 7.36 t5H, s, OC02CH2Ph), 6.81 ~lH, s, 5-H), 6.50 (lH, s, 8-H), 6.25 (2H, s, 2'- and 6'-H), 5.94 (2H, br.s, O-CH2-O), 5.23 (2H, s, -OC02CH2Ph), 4-~9 (1~, d, J=4 Hz, 4-H), 3.66 (6H, s, 3',5'-OCH3), 2.8-3.0 (2H, m, 2",3"-OH,D20 exchanged), 1.38 (3H, d, J=5 Hz, ~"--CH3).
Example 1 Preparation of 2",3"-di-O-acetyl etoposide (VIII, R =CH3, R2=H, R =R4=CH3Co-) (â ) P~epar on o~ ~'-0-~en~y o~.ycarbo..~e~o~os de di-O-ace_atc ~ cetic anhydridc ~' ml) was adde_ _o a sclution of 4'-CBZ-etoposide (500 m~" O.69 mmol) ir. ?yric'ine (10 ~r,l) and thG mixt~are ~as st ~~~ed fo~ 4 ho~lrs at -oom tem?cra ure.
1 ~
~ 5-7 The reaction mixture wa~ q~lenched with methanol (2 ml), diluted with dichloromethane, and then washed successively with water, 5% HCl, and water. The organic layer was dried over sodium sulfate, filtered, and the filtrate :oncentrated in vacuo to obtain a colorless solid (551 mg, 99%).
Recrystallization of the crude solid from methanol gave 4'-0-benzyloxycarbonlyeto~oside 2",3"-di-0-acetate as colorless crystals (lst. 395 mg, 2nd. 60 m~).
MP 236-238C.
IR vma~ Pr) cm 1 3500(br), 1770, 1610.
UV ~max (~JeH) nm () 290 (3950).
Anal. Calcd- for C41H4217 C 61.04, H 5.25.
Found: C 60.64, H 5.36.
(b) Preparation of 2'',3''-di-O-AcetYl etoposide A stirred solution of the product of step (a) (405 mg, 0.5 mmol) in ethanol-acetone (4:', 15 ml) was hydrogenatec for 1.5 hr in the presence of 10,~ Pd-C (400 mg) at 1 a~m, and then the catalyst was filtered off. The filtrate w~s concentrated in vacuo ~.o obtain colorless solid (360 mg, ca.
100%), which was recrystallized from ethyl acetate-methanol to give the title com2ound (20s mg) as -olorless crystals.
Estimated purity: a0% (by HPLC).
MP 287-289C.
I~ v (K~r) cm 3500(b^), ;760, 1610.
UV ~m ; (MeOH) nm ( ) 2a0(sh, 12780~, 285 (4060).
~nai. Calcd. for C33H35O15: C 58.93, H 5.39.
Found: C 58.56, H 5.al.
l8~36~
ExamPle 2 Preparation e' *'-O-benzyloxyc~arbonyletoposide 2"-o-acetate and 3"-0-acetate Acetic anhydride (20 ~11, 0.18 mmol) and 4-dimethylaminopyridine (5 mg) were added to a solution of 4'-CBZ etoposide (142 mg, 0.2 mmol) in pyridine (8 ml) kept at -10C. The mixture was stirred for 30 min. at -10C, after which time additional acetic anhydI~ide (20 lJl, 0.18 mmol) was added. The reaction mixture was skirred for a further 30 min. at -10C, and then diluted with dichloromethane, and washed s~lccessively with ~ater, r % HCl, aq. sodium bicarbonate, and water. The organic phase was dried over ~'a2S04, filtered, and tlle filtrate was concentrated in vacuo to afford colorless powder (117 mg) containing four components (silica gel TLC: Rf 0.56, 0.46, 0.34 and 0.24; hexane/acetone = 1/1). The mixture was separated by silica gel column chromatography (2.5%
met`~nanol-dichloromethan). Fractions showing a spot of Rf O.56 were combined and evaporated _n v_cuo to c~-ve 1 m~ (1%) of 2",3"-di-0-acetyl-4'-0-benzyloxycarbonyletoposide(4).
Simiiarly, fractions of Rf 0.46, 0.34 and 0.24 2fforded 3"-0-acetyl-4'-0-benzylo~;ycarbonyletoposide(3), 2"-0-ace-yl-4'-0-benzyloxycarbor..yletoposide(2) and 4'-CBZ etoposide (1, 60 mg, 42%) as colorless powder, respec~i.vely. Mixtu;es of 3 and 4 (18 mg), 2 and 3 (15 mg), and 1 and 2 (6 mg) were also obtained from the in-between fractions.
2"-0- cet;l-4'-0-be~ loxv arbol~yletoposide MP 224-227 '.
IR ''ma~ (KB~) cm 3400(br), 1740, 1600.
UV ~ (MeOH) nm () 2al (4200).
Anal Calcd for C39H4016-1/2H2 C 61.25, E1 5-~7 Found: C 60.84, H 5.25.
~n~3067 -`-3"-0-acetyl-4'-0-benzyloxycarbonyletoposide MP 139-145C.
IR vmax (KBr) cm 1 3500(br), 1770, 1600.
UV ~m x (MeOH) nm (E) 291 (3920).
Anal. Calcd for C39H40016 1/2H2 C 60.54~ H 5.34-Found: C 60.27, H 5 26.
Example 3 Preparation of 2"-0-acetyl etoposide (VIII, R1=CH3, R2=H, R3=CH3Co-, R4=H) The procedure described in Example 1, step (~) was followed using 2"-0-acetyl-4'-0-benzyloxycarbonyletoposide (25 mg, 0.03 mm^l) to give the title compound (21 mg, ca.
100%) as colorless powder. Estimated purity 90% (by HPLC).
MP 144-147C.
IR vmax (KBr) cm 3450(br), 1770, 1740, 1610.
UV ~max (MeOH) nm (E) 240(sh 12900), 284 (4030).
Anal- Calcd- for C31H3414 C 59.04, H 5.43.
Found: C 58.65, H 5.46.
ExamPle 4 Preparation of 3"-0-acetyl-etoposide (~7III, R1=
CH3, R =H, R =H, R -CH3CO) The procedure described in Example 1, ~tep (b) was followed using 3"-0-acetyl-4'-0-benzyloxycarbonyletoposide (10 mg, 0.01 mmol) to give the title cotnpound (9 mg, ca.
100%) as a coiorless powder. Estimated purity: 85o (b~
H?LC).
%0~ )6'7 - ~
MP 22~ 2~8C
IR ~ma~ (KB~) cm 1 3450(br), 1760, 1610.
UV ~max (MeOH) nm (~) 240(sh, 12610), 285 (3960).
Anal- Calcd. for C~lH3*14 1/2H2C~ C 5~.21~ H 5-52-Found: C 58.22, H 5.42.
Example 5 Preparation of 2"-O-acetyletoposide and 3"~0-acetyletoposide from a mixture of 2 and 3 A stirred solution of a mixture of 2 and 3 (60 mg, 0.08 mmol) in ethanol-acetone (4:1, 5 ml) was nydrogenated for 1.5 hr in the presence of 10% Pd-C (30 mg) at 1 atm, and then the catalyst was filtered off. The iltrate was concentrated to give a miY.ture of regio isomers t50 mg, ca.
100%), which was separated by silica gel column to give 2"-0-acetyletoposide (10 mg, 20%), 3"-0-acetyletoposide (4 mg, 8%), and a mixture of them (30 mg, 61%) as colorless powder.
Example 6. Preparation of 2",3"-di-G-formyl etoposide (VIII; Rl=CH3, R2=H, R3=R =HCO-) a. Preparation of 4'-0-benzyloxycarbony~eto~?oside 2'' 3''-di-O-formate To a solution of 4'-CBZ etoposide (].00 mg, 0.14 mmol) in pyridine (0.7 ml) was added dropwise a mixture of 99%
formic acid (1.4 ml) and acetic anhydride (0.56 ml) a~ 0C.
The reac ion mixture was stirred at room temperatilre for `
hours and then diluted with dichloromethane, and washed with water. The organic phase W25 drie~ o~er '~1a2SO4, then filtered off, and the filtrate was concentrated in vacuo to obtain colo-less solid (122 mg), which was purified by 80~7 -`~
silica ~1 column (MeOil:CH2Cl~ = 1:50) to give 4'-0-benzyloxycarbonyletoposide 2"-3"-di-0-formate (91 mg, 84%~ as amorphos powder.
H NMR (CDC13) ~ 8.04 & 7.80 (each lH, s, CHO), 7.36 (5H, s, PhCH2CO), 6.72 (lH, s, H-5), 6.53 (lH), s, H-8), 6.23 (2H, s, 2',6'-H), 5.97 (2H, s, OCH20), 5.23 (2H, s, PhCH2)CO), 5.4-4.0 (9H, m), 3.66 (6H, s, 3',5'-OCH3~, 3.6-2.6 (6H, m), 1.35 (3H, d, J=5 H-, 8"-CH3).
b. PreParation of 2",3"-~i-0-formyletoposide A stirred solut~on of the product of step (a) (71 m~, 0.009 mmol) in ethanol-acetone (4:1, 2.5 ml) was hydro~enated for 1.5 hr in the presence of 10% Pd-C (50 mg) at 1 atm. The catalys' was filtered off and the filtrate was concentrated in vacuo to give the title com~ound as colorless powder (59 mgj ca. 100~). Estimated purity 80% by HPLC.
MP 7-280C.
IR vmax (KBr) cm 3400, 1760, 1740, 1610 UV ~max (MeOH) nm ~) 236 (sh, 14,100~, 285 (4,230) H NMR (CDC13) ~ 8-04 & 7.83 (each lH, s, CHO), 6.72 (lH,s,H-5), 6.53 (lH, s, H-8), 6.23 (2H, s, 2',6'-H), 5.98 (2H, ~, OCH20), 5.6-4.1 (9H, m), 3.76 (6H, s, 3',5'-OCH3), j.7-2.6 (6H, m), 1.35 (3~, d, J=5 H~, 8"-CH3).
Ana' Calc~ f~r C31H3215 C 57-26~ H 5.0C.
, OU.IQ: C 57 . C~ 5 . 00 xam~le 7. Preparation o~ ben~yioxycarbonyle'-oposi~e 2"-0-benzoate, 3"-ben_oate, an~ 2",3"-di-C-ben7oate -~ Z0~89~7 ~
Benzoyl chloride (100 ~l, 0.86 mmol) was added to a solution of 4'-0-CBZ etoposide (500 mg, 0.7 mmol) in pyridine (5 ml). The reaction mixture was stirred at room temperature for 3 days, diluted with dichloromethane, and then washed successively with water, 5% HCl, and water. The organic layer was dried over Na2SO4, filtered, and the filtrate was concentrated in vacuo to give pale yellow oil, which contained three new components (silia gel TLC: Rf 0.50, 03.8 and 0.21; dichloromethane/methanol = 100/1). he mixture was separated by silica gel col~imn chrom~tography (40% hexane-acetone) to obtain 2",3"-di-0-benzoate 7 ~Rf;
0 50, 30 mg, 5%), 3"-0-benzoate 6 (Rf; 0.38, 227 mg, 40%), 2"-0-benzoate 5 (Rf; 0.21, 70 mg, 12%) and recovered 4'-CBZ
etoposide 1 (215 mg, 43~).
4'-BenzyloxycarbonYletoposide 2"-0-benzoate (5) IR vmax (~Br) cm 3400(br), 1700, 1720, 1600 UV ~ ax (MeOH) nm (~) 281 (4,050) H NMR (CDCl3) ~ 7.3-7.9 (10H, m, 2xC5H5, 5-17 (lH, t, J=8 Hz, 2"-H0, 5.01 (lH, d, J=8 Hz, 1"-H , 4.77 (lH, q, J=5 Hz, 7"-H), 3.2-3.8 (lH, m, 3"-H), 2.75 (iH, br, 3"-OH), 1.31 (3H, d, J=5 Hz, 7"-CH3).
Anal Calcd for C44H4216 C 63-92~ H 5-12-Found: C 63.80, H 5.17.
4'-0-Benzyloxycarbonyletoposide 3" O-b~nzoate (6) IR vmax (~r) cm 3500(br), 1770, 1600 max (MeO~) nm ( E ) 225 (ch 30 390) 281 H NMR (CDCl3) ~ 7.4-8.0 (10H, m, 2xC6H5, 5.34 (lH, t, J=9 Hz, 3"-H, 4.74 (lH, d, J=8 Hz, 1"-H), 4.68 (;H, q, J=5 Hz, 7"-H), 3.60 (lH, m, 2"-H), 1.28 (3H, d, J=5 Hz, 7"-CH_).
L8~)67 Anal Calcd for C44H4216 C 62-55~ H 5-25-Found: C 62.73, H 5.29.
4'-0-Ben2yloxycarbonyletoposide 2'',3''-di-O-benzoate (7j IR vmax (KBr) cm 1770, 1730, 1600 UV ~max (MeOH) nm (E) 227 (sh, 40,270), 282 (4,550) lH NMR (CDC13) ~ 7.3-7.9 (lOH, m, 2xC6H5), 5.55 (lH, t, J=8 Hz, 3"-H), 5.32 (lH, t, J=8 Hz, 2"-~:, 4.93 (].H, d, J=8 H~, l"-H), 4.50 (lH, q, 7"-H), 1.31 (3H, d, J=5 Hz, 7"-CH3).
Anal Calcd for C51H4617 C 65-80~ H 4-55-Found: C 65.28, H 4.98.
Example 8. Preparation of 2"-0-benzoyletoposide (VIII, Rl=CH3, R2=H, R3=PhCO-, R4=H) A stirred solution of 4'-benzyloxycarbonyletoposide 2"-O-benzoate (54 mg, 0.07 mmol) in ethanol-acetone (2:1, 3 ml) was hydrogen~ted for 2 hours i.n the presenc~ of 10%
palladium on carbon (50 mg) at 1 atm. The catalyst was filtered off and the filtrate was concentrated to give the title compound (40 mg, 88% as colorless powder. Estimated purity 95% by HPLC.
MP ~89-292C.
IR ~max (KBr) cm 3450, 1760, 1730 UV ~max (MeOH) nm (~) 283 (4,470) H NMR (CDC13) ~ 7.4-7.8 (5H, m, COC6H5), 5 07 (1~, dd, J=7.7 & 9.2 Hz, 2"-H), 4.89 (lH, d, J=7 7 Hz, l"-H), 4.80 (lH, q, J=5.1 Hz, 7"-H), 4.23 (1~, dd, J=4.8 & 10.3 Hz, 6"-Heq), 3.98 (lH, dt, J=3.3 & 9.2 Hz, 3"-H), 3.64 (lH, t, J=10.3 Hz, 6"-Hax), 3.47 ~lH, t, J=9.2 .~z, 4"-H), 3.40 (lH, 8~Çi7 ''' dt, J=4.8 ~ 10 Hz, 5"-H), 2.74 (lH, d, J=3.3 Hz, 3"=OH), 1.41 (3H, d, J=5.1 Hz, 7"-CH3).
i~nal Calcd for C36H36Ql4 H20 Found: C 61.10, H 5.40.
Example 9. Preparation of 3"-0 Benzoyletoposide (VIII, R1=
CH3, R2=H, R3=H, R4=PhCO) According to the procedure of Example 8, 34 mg (C.04 mmol~ of 4'-0-benzyloxycarbonyletoposide 3"-0-benzoate was hydrogenated to give the title compound (27 mg, 95%) as colorless solid, which was purified on column chromatography to give colorless crystals. Estimated purity 95% by HPLC.
MP 188-191C.
IR `'max (KBr) cm 1 3400 (br), 1730.
UV ~max (MeOH) nm (E) ~23 (35,900), 280 (4,030).
H NMR (CDCl3) ~ 7.4-7.8 (5H, m, COPh), 5.38 ~lH, t, J=9.2 Hz, 3"-H), 4.79 (lH, d, J=7.7 Hz, l"-H), 4.72 (lH, q, J=5.1 Hz, 7"-H), 4.22 (lH, dd, J=5 & 10 Hz, 6"-Heq), 3.67 (lH, brt, J=8 Hz, 2"-H;, 3.6~ (lH, t, J=10 Hz, 6"-H~ax?~ 3.59 (lH, t, J=9.4 Hz, 4"-H), 3.47 (lH, m, 5"-H), 2..58 (lH, br, 2"-OH), 1.31 (3H, d, J=5.1 Hz, 7"-CH3).
Anal Calcd for C36H36O14 2H2O
Found: C 59.14, H 5.10.
Exam~le 10. Preparation of 2",3"-O-di-benzoyletoposide (VIII, R1=CH3, R2=H, R3=R4=PhCO-) A stirred solution of 4'-0-benzyloxycârbonyletoposide 2",3"-di-0-benzoate (130 mg, 0.14 mmol) in ethyl acetate-ethanol-acetone (4:4:1, 4.5 ml) was hydrogenated according to the procedu e of Example 8 to give the title compound (112 mg, ca. 100%) as colorless powder. E~'imateci ~0~ 67 purity 90% by HPLC.
MP >295C.
IR vmax (KBr) cm 1 1770, 1730.
UV ~max (MeOH) nm (E) 226 (sh, 24,200), 281, (3,470).
H NMR (CDCl3) ~ 7.3-8.0 (lOH, m, 2xCOC6H5), 5.67 (lH, t, J=9.5 Hz, 3"-H), 5.35 (lH, dd, J=8.1 & 9.7 Hz, 2"-H), 498 (lH, d, ~=8.1 Hz, l"-H), 4.73 (lH, q, J=4.8 Hz, 7"-H), 4.28 (lH, dd, J=4.4 & 10.3 Hz, 6"-Heq), 3.69 (lH, t, J=9.5 Hz, 4"-H), 3.67 (lH, t, J=10 Hz, 5"-Hax), 3.56 (lH, dt, J=9.5 &
lO H~, 5"-H), 1.33 (3H, d, J=4.8 Hz, 7"-CH3).
Anal Calcd for C43H400l5 H2 Found: C 62.99, H 4.99.
Example 11 The general procedure described in Example 1 is follo~!ed using 4'-CBZ teniposide instead of 4'-CBZ etoposide to provide 2",3"-di-0-acetyl teniposide.
Example 12 The general procedure described in Examples 2-4 is repeated using 4'-CBZ teniposide instead of 4'-CBZ etoposide to provide 2"-0-acetyl teniposide and 3"-0-teniposide.
Example 13 The gene-21 proces~lre s;essribed in ~x~.mF`e ~ i.s repeated usiilg ~'-C~Z teni?^s_dP insteas~ of ~ -C-Z etopos de ~o pro~.-ide 2",3"-di-0-formyl teniposide.
- 2~
ExamPle 14 The general procedure of Examples 7-10 is repeated using 4'-CBZ teniposide instead of 4'-CBZ etoposide to provide 2"-0-benzoyl teniposide, 3"-0-benzoyl teniposide and 2",3"-di-0-benzoyl teniposide.
CH~ ~
~ H ~..
~0 <~ ~"'\~
CU3~0CU3 ~ ~oeos!de_ehoseha ~e SUMM~R~' OF TEE INV~NTION
The present invention provides compounds having the formula (VIII) R L~
(~ ~0 CH~ CH3 ORS
~Vl I I ) wherein R2 is H and Rl is selected fro~ the group consisting f (Cl-lo)alkyl~ (C2-lO)alkenylr ~C5 6)cycloalkyl~ 2-furyl, y , ( 6_l0)aryl, and (C7_14)aralkyl; or R and ~ a~-e each (Cl lO)alkyl; or Rl, R and the carhon to which they are at_ached together represent ~C5 6)cyclo21kyl; one of R
and R^ is ~ and the other is selected from the group 2~ 6~ ~
consisting of (Cl 5)alkanoyl and benzoyl; or R3 and R4 are the same and are selected from the group consisting of (C1 5) alkanoyl and benzoyl; R5 is H or a phosphate group.
Another aspect of the present invention provides pharmaceutical compositions comprising an antitumor effective amount of a compound of formula (VIII) and a pharmaceutically acceptable carrier.
Yet a further aspect of the present invention provides a method for inhibiting tumor growth in a tumor bearing mammalian host which comprises administering to said hcst an antitumor amount of a compound of form~lla (VIII).
DET~ILED DESCr~IPTION OF `1~ INVENTION
A preferred embodiment of compounds of formula (VIII) comprises those compounds wherein R2 is H and ~1 is selected from the group consisting of methyl, 2-thienyl, and phenyl, with methyl being the most pleferred substituent.
A further preferred embodiment pro~i.des _ompounds of formula (VIII) wherein R3 and R4 are each ~C1 5~alkanoyl; a most preferred embodiment provides compounds of formula (VIII) wherein R3 and R4 are each a formyl gro~lp.
Yet a further preferred embodiment pro~,ides compounds of formula (VIII) where-in one of h- and R4 is H, and the other is (Cl 5~alkanoyl or benzoyl.
As used herein, the term "phosphate".ir.~ludes the-group-PO~H2 and pharmaceutically acceptable salts tilereof.
Z()1~7 -~
Pharmaceutica'1y acceptable salts include both monobasic and dibasic salts wherein the cation includes, but not limited to, alkali metals, e.g. sodium, potassium and lithium;
alkaline earth metals, e.g. magnesium, calcium and barium;
orgnic amine salts, e.g. ammonium.
The novel compounds of the present invention may be prepared by reactin~ a 4'-phenol protected 4'-demethylepipodophyllotoxin g]ucoside o formula ( X~) with the desired carboxylic acid or an acylating equivalent thereof, followed by removal of the phenol protecting group.
R ~1~\ D
<~0 CH3 ~OCH3 y ~ a: Y ~ o ~ l i ng gl- oup lXb: Y = H
~ he phenol protecting group is not par~icularly restricted and may be any that can be irtroduced and removed with little or no adverse effect on other parts of the molecule. More particularly, the phenol protecting g~oup should be removable by methods that ~c not distu-b the acyloxy functionalities cn the sugar moiety o- the final produc~s. Methods fcr phenol protection a~-e well kno~r. in the art and as examples of aeneral classes of phenol protecting groups, mention can be made o~ ethers, acetzls, esters, and carbonatec. For the purpose of the present invention, we found the bPn~yloxycarbonyl racliczl mzy be 6~ ~, conveni~r_ly so employed. Thus by reacting a 4'-demethyl-epipodophyllotoxin glucoside of formula (IXb) with benzylchloroformate at low temperature, e.g. -15C, in an inert organic solvent such as methylene chloride and in the presence of a tertiary organic base such as pyridine, the corresponding 4'-0-benzylo~ycarbonyl-4'-demethylepipodo-phyllotoxin glucoside is obtained. Compounds of formula (IXb) and their synthesis are disclosed in US Patent No.
3,524,844.
The acylation of the sugar llydro~yl groups of compounds of formula (IXa) may be effected using the desired carboxylic acid or an acylating agent derived therefrom, for example an acid halide such as acid chloride, active ester derived from N-hydroxysuccinimide or 1-hydroxytriazole, and symmetrical or mixed anhydride. When the carboxylic acid is used as the acylating species, it is preferably used in conjunction with a condensing agent for e~ample a carbodiimide such as dicyclohexylcarbodiimide (DCC).
Acylating 2gents are C1 5alkanoic acids and ber.~oic acid and acylating equivalents thereof as above d~scribed. Alkanoic acids include, but are not limited to, formic acid, acetlc acid, propionic acid, butyrlc acid, pentanoic acid, and branched alkanoic acids such as 2-methylpropionic acid and 3-methylb~_tanoic acid.
The acylation reaction may be carried out in an inert organic solvent such as pyridine, and prefera~ly in-lude n the reaction mixture an acid acce?~o~ ~hen acid is ex?ec~ed to be a by-?roduct; suitable acia acceptors are for example tertia-y amine bases such as pyri-ine, triethylamine, diisopropylethylamine and the like, or inorganic bases such as sodium anG potassium carbonates. The reaction may be conGucted a~ tempera~ures conduci~-e to forma_ion of the Z5)~6~ -~
~sired products an~ may be from about -15C to about 50C;
the reaction may ta~e several minutes to several days foi-completion deppending on the nature of the reactants and choice of reaction conditions such as temperature.
The acylation is not regioselective and may result in a mixture of 2"-monoacylated product, 3"-monoacylated product and 2",3"-bisacylated product. However, when at least two equivalents of the acyla,ing agent is used relative to the epipodophyllotoxin reactant, the bis-acylated derivative is produced predominantly. The proportion of monoacylated product and bisacylated product may be controlled to some extent by manipulative reaction parameters according to principles commonly practiced in the art, e.g. varying the relative amounts of reactants, or varying reaction conditions such as temperature and length of reaction time.
In general when equimolar of the reactants are e~..loyed, reduced reaction temperature and shorter reaction time tend to increase the amount of monoacylated products formec.
Alternatively, monoacylated products may be obtained by first protecting one of the sugar hydroxyl groups of the 4'-phenol protected epipodophylloto~in glucoside starti;.g material. The protecting group for the sugar hydroxyl group is preferably a bul~y protecting group, for example the t-butyldimethylsilyl group, in order to minimize derivatizing both hydroxyl groups. Again, protection of the sugar hydroxy-l groups is not regioselective; however, if desi-ed, the 2"-protec'ed cr 3"-prot~_ted compounds are easily separa~ed chroma.og aphically. We have carried ou formylation reactions of eto?oside; howe~er, we have no~
been slccessful in isola~ing either of ~he expected mono ormyl derivatives of etoposide.
Z01~3~6~7 The product mixture comprising pl~enol-protected acylated derivatives may be separated into individual components using conventional separat;on techniques such as column chromatography. Deprotection of the phenolic hydroxyl group and, if necessary, the sugar hydroxyl gro-lp provides compounds of the present invention. The deprotection may be effected prior to or subsequent to the isolation of the individual components; the order is not critical. The protecting group i~ removed by usin~ methods suitable for the protecting group selected and these are generally well known in the art. For example, when the protecting group is benzyloxycarbonyl, it may be removed by catalytic hydrogenation at atmospheric precsure using palladium on carbon as the catalyst; the t-butyldimethyl-silyl group may be removed by heating in an aqueous alcoholic solution or by treating with fluoro anion or aqueous acetic acid.
Following the removal of the phencl protecting group, the ~'-phenol g~oup may be further derivati~ed to provide the 4'-phosphate of formula (VIIl). Phosphorylation of the 4'-phenol may be carried out using conventional methods.
For example, a compound of formula (VIII) wherein R5 is H
may be reacted with a phosphorylating agent such as phosphorous o~ychloride; the phosphoryl chloride intermediate i~ hydrolyzed i_ situ to give the 4'-pllosphate.
Hydrolysis in the presence of a base e.g. sodiu~ bicarbonate provides the corresponding salt. This procedure as well as other potentially applicable phosphorylation methods is disclosed in U.S. patent application GB2,207,674 which is hereby incorporated by refe ence.
201~3~6'7 It is to be understood that synthesis of compounds of the present invention is not limited to the procedures and reagents outlined above, but may include other methods capa~le of acylating the hydroxyl groups on the sugar portion of 4'-demethylepipodophyllotoxin glucosides. The reaction conditions will of course vary with the choice of startlng materials but may be ascertained by a skilled artisan without und~le experimentation.
BIOLOGICAL ACTIVITY
Representat-ive compounds of the present invention were evaluated for antitumor activity against murine transplantable P38~ le~kemia. Female CDF1 mice were inoculated intraperitoneally with 0.4 ml of diluted ascitic fluid containing 106 lymphocytic leukemia P388 cells. Test compounds were administered intraperitoneally as a .single dose on day 1 and animals were observed for 50 days. The percent increase of median survival time (MST) of treated animals over that of un~reated control animals was determined and re?orted as ~O T/C. Compounds sho-;ing % T/C
values of 125 or greater are considered to have significant antitumor activity. Table I presents the results of the in vivo evaluation; only th~ maximum ,O T~C and the dose giving the maximum effects are reported.
~O~ 7 Table I. Antitumor activitv _f~a1nst P3~8 leukemia Compound _o e_~mg~kgf/day~ ~T/C MST
Example 1 30 165 Example 3 30 240 Ex~mple 4 30 210 Etoposide 30 182 Example 6 120 >500 ~3/4)*
Example 8 120 14r`
Example 9 60 f~20 ( l/f~ ) Example 10 120 125 Etoposide 120 >390 (6/12) *Number of survivors/tested on day S0 ~/~ /S/
In vitro cytotoxicity of compounds of the presen-t R G invention was evaluated against ' ]~r c~ ' l r~ r ~f~ B16-F10 murine melanoma ~ oser human colcn carcinoma~
bl lT '~ ~ ~;r r ~ r r r ~ ~ ~ ~ I~r~ ~ ^ T~
r^r- ~t~ -1 i nf~ ~ - r~ r ~~ L - r~
Iql; E R/6~ o f~L Mo~J
n~(f -~ I~C~ ~f ~xponentially gr ~ e harvested, counted and suspended in the culture me~dium at the ~; concentrations of 1.5 x 104 and 3 x 104 cells/ml, respectively. Twenty-four hours after p].anting c~11 suspensicn (î80 mcl) into wells of a rJo-weil microti~er plate, test materia s (20 mcl) were added to the wells and the ?lates were incubated for /2 hours. The cyto oxi_ activi~ies against ~he tum~r cells were colorimet~-ically 2(~ 7 determined at 540 nm after staining viable cells with ~/~ ~&~ neutral red solution. ~or cytotol~ic-it~ pcrimcn~-~
R ~ K56. ~nd ~56~ DM ccllc, ~00 m~l-ef thc ccll ~u3~cnsi~
~,~k,~ 104 ccll-~ml) ~ac incub~ted ~.~ith tcct m~e~al3 ~100 mel) a' 37C, 5,~ C~2 for 4~ hollr~ in - 4 ~cll tis~uc cult~e-p~
~hc c~toto~ic actir'~ Jas-~e~crmincd b~ e~unt~ e num~cr of colls uEing a cell ~ ntcr (s~Gme~ cc 120.~). The results are summarized in Table II.
~/z6/~/ u~Y~ ~ n~ ~JQ~
Table II. In vitro cytotoxicity_~a_nst ~ariouL ce'~ lin~
h~
~l');
IC50 (mcg~
Compound B16-F10 Moser~ I~rC ,'~
Example 3 1.6 1.1 0.
~/6/~ Example 4 1.4 ND* ~D ~
Example l 1.3 <0.8-0.1~ ~ ' `50 Example 8 0.92 ND~D ,~\ ~ID
Example 9 0.17 0.83O~4~J/~ \\ 6.7 Example lO 4.4 ND I~
Example 6 2.0 0.84 Etoposide 0.45 ND 'O;O
ND: Not determined Accordingly, the present invention provides a method ror inhibiting mammalian tumors ~hich comprises aaministering an effective tumor-inhibiting dose of an antitumor co~pound of formula VIII to a tumor bearing host.
For this purpose, the druc may be admin,stered by conventional routes including, but not limited to, intravenous, intramuscular, intratumoral, intraarterial, ~8~67 ^ ~
intralymphc~ic, and oral.
A further aspect of the present invention provides a pharm2-eutical composition whi.ch comprises a compound of formula VIII and a pharmaceutically acceptable carrier. The antitumor composition may be made up of any pharmac~-~ti-cal form appropriate for the desired route of administra-tion. Examples of such compositions include solid compositions fGr oral administration such as tablets, capsules, pills, powders and granules, liquid compositlons for oral administration such as solutions, suspensions, syrups or elixirs and preparations for parenteral administration such as sterile solutions, suspensions or emulsions. They may also be manufactured in the form of sterile solid compositions which can be dissolved in sterile water, physiological saline or some other sterile injectable medium immediately before use.
Optimal dosages and regimens for a givPn mammalian host can be readily ascertained by those skilled n the art. It will, of co~rse, be ap~reciated that the actual dose used will vary according to the particu].ar co.m?osition formulated, the partic~lar compound used, the mode of application and the particular site, host and disease being treated. Many factors that modify the action of the drug will be taken into account incl~lding age, weight, sex, diet, time of adm.inistration, rou~e of administra~ion, rate o.
e~cretion, condition of the patien', drug combinatiolls, reaction sensiti~;ities and sP~erity of the disease.
The following exam?les a-e for illus~rative pu-?oses only and should not be construPd as limiting the _cope of the invention which is defined solely by the Claims appended to the specification.
,;~ a67 Preparation of 4'-O-Benzyloxycarbonyl Etoposide (4'-CBZ
etoposide) Benz~ lchloroformate (1.98 ml, 15 mmol) was addeP over a period of 30 minutes to a mixture of etoposide (5.88 g, 10 mmol) and p~,ridine (10 ml) in methylene chloride (100 ml) kepl; at -15C. The mixture was stirred at -15C for an additional ho~lr, washed successively ~ith 5% HCl, aqueous sodium hicarbonate, and water, and then driec o~er anhydrGus sodium sulfate. The solvent was evapora1:ed to gi~e 8.05 ~3 of the crude product which ~,~as purified on a silica gel column (5% methanofl-methylene chloride) to give 6.93 g (96%) of 4'-O-benzyloxycarbonyl etoposide as a colorless semi-solid. M.P. 152-155C
IR ~max (Nujol) cm : 320"0-3600 (OH), 1760 (lactone &
4'-0-benzyloxy carbonyl), 1600 (aromatic).
H NMR (60 MHz, CDCl_) ~ 7.36 t5H, s, OC02CH2Ph), 6.81 ~lH, s, 5-H), 6.50 (lH, s, 8-H), 6.25 (2H, s, 2'- and 6'-H), 5.94 (2H, br.s, O-CH2-O), 5.23 (2H, s, -OC02CH2Ph), 4-~9 (1~, d, J=4 Hz, 4-H), 3.66 (6H, s, 3',5'-OCH3), 2.8-3.0 (2H, m, 2",3"-OH,D20 exchanged), 1.38 (3H, d, J=5 Hz, ~"--CH3).
Example 1 Preparation of 2",3"-di-O-acetyl etoposide (VIII, R =CH3, R2=H, R =R4=CH3Co-) (â ) P~epar on o~ ~'-0-~en~y o~.ycarbo..~e~o~os de di-O-ace_atc ~ cetic anhydridc ~' ml) was adde_ _o a sclution of 4'-CBZ-etoposide (500 m~" O.69 mmol) ir. ?yric'ine (10 ~r,l) and thG mixt~are ~as st ~~~ed fo~ 4 ho~lrs at -oom tem?cra ure.
1 ~
~ 5-7 The reaction mixture wa~ q~lenched with methanol (2 ml), diluted with dichloromethane, and then washed successively with water, 5% HCl, and water. The organic layer was dried over sodium sulfate, filtered, and the filtrate :oncentrated in vacuo to obtain a colorless solid (551 mg, 99%).
Recrystallization of the crude solid from methanol gave 4'-0-benzyloxycarbonlyeto~oside 2",3"-di-0-acetate as colorless crystals (lst. 395 mg, 2nd. 60 m~).
MP 236-238C.
IR vma~ Pr) cm 1 3500(br), 1770, 1610.
UV ~max (~JeH) nm () 290 (3950).
Anal. Calcd- for C41H4217 C 61.04, H 5.25.
Found: C 60.64, H 5.36.
(b) Preparation of 2'',3''-di-O-AcetYl etoposide A stirred solution of the product of step (a) (405 mg, 0.5 mmol) in ethanol-acetone (4:', 15 ml) was hydrogenatec for 1.5 hr in the presence of 10,~ Pd-C (400 mg) at 1 a~m, and then the catalyst was filtered off. The filtrate w~s concentrated in vacuo ~.o obtain colorless solid (360 mg, ca.
100%), which was recrystallized from ethyl acetate-methanol to give the title com2ound (20s mg) as -olorless crystals.
Estimated purity: a0% (by HPLC).
MP 287-289C.
I~ v (K~r) cm 3500(b^), ;760, 1610.
UV ~m ; (MeOH) nm ( ) 2a0(sh, 12780~, 285 (4060).
~nai. Calcd. for C33H35O15: C 58.93, H 5.39.
Found: C 58.56, H 5.al.
l8~36~
ExamPle 2 Preparation e' *'-O-benzyloxyc~arbonyletoposide 2"-o-acetate and 3"-0-acetate Acetic anhydride (20 ~11, 0.18 mmol) and 4-dimethylaminopyridine (5 mg) were added to a solution of 4'-CBZ etoposide (142 mg, 0.2 mmol) in pyridine (8 ml) kept at -10C. The mixture was stirred for 30 min. at -10C, after which time additional acetic anhydI~ide (20 lJl, 0.18 mmol) was added. The reaction mixture was skirred for a further 30 min. at -10C, and then diluted with dichloromethane, and washed s~lccessively with ~ater, r % HCl, aq. sodium bicarbonate, and water. The organic phase was dried over ~'a2S04, filtered, and tlle filtrate was concentrated in vacuo to afford colorless powder (117 mg) containing four components (silica gel TLC: Rf 0.56, 0.46, 0.34 and 0.24; hexane/acetone = 1/1). The mixture was separated by silica gel column chromatography (2.5%
met`~nanol-dichloromethan). Fractions showing a spot of Rf O.56 were combined and evaporated _n v_cuo to c~-ve 1 m~ (1%) of 2",3"-di-0-acetyl-4'-0-benzyloxycarbonyletoposide(4).
Simiiarly, fractions of Rf 0.46, 0.34 and 0.24 2fforded 3"-0-acetyl-4'-0-benzylo~;ycarbonyletoposide(3), 2"-0-ace-yl-4'-0-benzyloxycarbor..yletoposide(2) and 4'-CBZ etoposide (1, 60 mg, 42%) as colorless powder, respec~i.vely. Mixtu;es of 3 and 4 (18 mg), 2 and 3 (15 mg), and 1 and 2 (6 mg) were also obtained from the in-between fractions.
2"-0- cet;l-4'-0-be~ loxv arbol~yletoposide MP 224-227 '.
IR ''ma~ (KB~) cm 3400(br), 1740, 1600.
UV ~ (MeOH) nm () 2al (4200).
Anal Calcd for C39H4016-1/2H2 C 61.25, E1 5-~7 Found: C 60.84, H 5.25.
~n~3067 -`-3"-0-acetyl-4'-0-benzyloxycarbonyletoposide MP 139-145C.
IR vmax (KBr) cm 1 3500(br), 1770, 1600.
UV ~m x (MeOH) nm (E) 291 (3920).
Anal. Calcd for C39H40016 1/2H2 C 60.54~ H 5.34-Found: C 60.27, H 5 26.
Example 3 Preparation of 2"-0-acetyl etoposide (VIII, R1=CH3, R2=H, R3=CH3Co-, R4=H) The procedure described in Example 1, step (~) was followed using 2"-0-acetyl-4'-0-benzyloxycarbonyletoposide (25 mg, 0.03 mm^l) to give the title compound (21 mg, ca.
100%) as colorless powder. Estimated purity 90% (by HPLC).
MP 144-147C.
IR vmax (KBr) cm 3450(br), 1770, 1740, 1610.
UV ~max (MeOH) nm (E) 240(sh 12900), 284 (4030).
Anal- Calcd- for C31H3414 C 59.04, H 5.43.
Found: C 58.65, H 5.46.
ExamPle 4 Preparation of 3"-0-acetyl-etoposide (~7III, R1=
CH3, R =H, R =H, R -CH3CO) The procedure described in Example 1, ~tep (b) was followed using 3"-0-acetyl-4'-0-benzyloxycarbonyletoposide (10 mg, 0.01 mmol) to give the title cotnpound (9 mg, ca.
100%) as a coiorless powder. Estimated purity: 85o (b~
H?LC).
%0~ )6'7 - ~
MP 22~ 2~8C
IR ~ma~ (KB~) cm 1 3450(br), 1760, 1610.
UV ~max (MeOH) nm (~) 240(sh, 12610), 285 (3960).
Anal- Calcd. for C~lH3*14 1/2H2C~ C 5~.21~ H 5-52-Found: C 58.22, H 5.42.
Example 5 Preparation of 2"-O-acetyletoposide and 3"~0-acetyletoposide from a mixture of 2 and 3 A stirred solution of a mixture of 2 and 3 (60 mg, 0.08 mmol) in ethanol-acetone (4:1, 5 ml) was nydrogenated for 1.5 hr in the presence of 10% Pd-C (30 mg) at 1 atm, and then the catalyst was filtered off. The iltrate was concentrated to give a miY.ture of regio isomers t50 mg, ca.
100%), which was separated by silica gel column to give 2"-0-acetyletoposide (10 mg, 20%), 3"-0-acetyletoposide (4 mg, 8%), and a mixture of them (30 mg, 61%) as colorless powder.
Example 6. Preparation of 2",3"-di-G-formyl etoposide (VIII; Rl=CH3, R2=H, R3=R =HCO-) a. Preparation of 4'-0-benzyloxycarbony~eto~?oside 2'' 3''-di-O-formate To a solution of 4'-CBZ etoposide (].00 mg, 0.14 mmol) in pyridine (0.7 ml) was added dropwise a mixture of 99%
formic acid (1.4 ml) and acetic anhydride (0.56 ml) a~ 0C.
The reac ion mixture was stirred at room temperatilre for `
hours and then diluted with dichloromethane, and washed with water. The organic phase W25 drie~ o~er '~1a2SO4, then filtered off, and the filtrate was concentrated in vacuo to obtain colo-less solid (122 mg), which was purified by 80~7 -`~
silica ~1 column (MeOil:CH2Cl~ = 1:50) to give 4'-0-benzyloxycarbonyletoposide 2"-3"-di-0-formate (91 mg, 84%~ as amorphos powder.
H NMR (CDC13) ~ 8.04 & 7.80 (each lH, s, CHO), 7.36 (5H, s, PhCH2CO), 6.72 (lH, s, H-5), 6.53 (lH), s, H-8), 6.23 (2H, s, 2',6'-H), 5.97 (2H, s, OCH20), 5.23 (2H, s, PhCH2)CO), 5.4-4.0 (9H, m), 3.66 (6H, s, 3',5'-OCH3~, 3.6-2.6 (6H, m), 1.35 (3H, d, J=5 H-, 8"-CH3).
b. PreParation of 2",3"-~i-0-formyletoposide A stirred solut~on of the product of step (a) (71 m~, 0.009 mmol) in ethanol-acetone (4:1, 2.5 ml) was hydro~enated for 1.5 hr in the presence of 10% Pd-C (50 mg) at 1 atm. The catalys' was filtered off and the filtrate was concentrated in vacuo to give the title com~ound as colorless powder (59 mgj ca. 100~). Estimated purity 80% by HPLC.
MP 7-280C.
IR vmax (KBr) cm 3400, 1760, 1740, 1610 UV ~max (MeOH) nm ~) 236 (sh, 14,100~, 285 (4,230) H NMR (CDC13) ~ 8-04 & 7.83 (each lH, s, CHO), 6.72 (lH,s,H-5), 6.53 (lH, s, H-8), 6.23 (2H, s, 2',6'-H), 5.98 (2H, ~, OCH20), 5.6-4.1 (9H, m), 3.76 (6H, s, 3',5'-OCH3), j.7-2.6 (6H, m), 1.35 (3~, d, J=5 H~, 8"-CH3).
Ana' Calc~ f~r C31H3215 C 57-26~ H 5.0C.
, OU.IQ: C 57 . C~ 5 . 00 xam~le 7. Preparation o~ ben~yioxycarbonyle'-oposi~e 2"-0-benzoate, 3"-ben_oate, an~ 2",3"-di-C-ben7oate -~ Z0~89~7 ~
Benzoyl chloride (100 ~l, 0.86 mmol) was added to a solution of 4'-0-CBZ etoposide (500 mg, 0.7 mmol) in pyridine (5 ml). The reaction mixture was stirred at room temperature for 3 days, diluted with dichloromethane, and then washed successively with water, 5% HCl, and water. The organic layer was dried over Na2SO4, filtered, and the filtrate was concentrated in vacuo to give pale yellow oil, which contained three new components (silia gel TLC: Rf 0.50, 03.8 and 0.21; dichloromethane/methanol = 100/1). he mixture was separated by silica gel col~imn chrom~tography (40% hexane-acetone) to obtain 2",3"-di-0-benzoate 7 ~Rf;
0 50, 30 mg, 5%), 3"-0-benzoate 6 (Rf; 0.38, 227 mg, 40%), 2"-0-benzoate 5 (Rf; 0.21, 70 mg, 12%) and recovered 4'-CBZ
etoposide 1 (215 mg, 43~).
4'-BenzyloxycarbonYletoposide 2"-0-benzoate (5) IR vmax (~Br) cm 3400(br), 1700, 1720, 1600 UV ~ ax (MeOH) nm (~) 281 (4,050) H NMR (CDCl3) ~ 7.3-7.9 (10H, m, 2xC5H5, 5-17 (lH, t, J=8 Hz, 2"-H0, 5.01 (lH, d, J=8 Hz, 1"-H , 4.77 (lH, q, J=5 Hz, 7"-H), 3.2-3.8 (lH, m, 3"-H), 2.75 (iH, br, 3"-OH), 1.31 (3H, d, J=5 Hz, 7"-CH3).
Anal Calcd for C44H4216 C 63-92~ H 5-12-Found: C 63.80, H 5.17.
4'-0-Benzyloxycarbonyletoposide 3" O-b~nzoate (6) IR vmax (~r) cm 3500(br), 1770, 1600 max (MeO~) nm ( E ) 225 (ch 30 390) 281 H NMR (CDCl3) ~ 7.4-8.0 (10H, m, 2xC6H5, 5.34 (lH, t, J=9 Hz, 3"-H, 4.74 (lH, d, J=8 Hz, 1"-H), 4.68 (;H, q, J=5 Hz, 7"-H), 3.60 (lH, m, 2"-H), 1.28 (3H, d, J=5 Hz, 7"-CH_).
L8~)67 Anal Calcd for C44H4216 C 62-55~ H 5-25-Found: C 62.73, H 5.29.
4'-0-Ben2yloxycarbonyletoposide 2'',3''-di-O-benzoate (7j IR vmax (KBr) cm 1770, 1730, 1600 UV ~max (MeOH) nm (E) 227 (sh, 40,270), 282 (4,550) lH NMR (CDC13) ~ 7.3-7.9 (lOH, m, 2xC6H5), 5.55 (lH, t, J=8 Hz, 3"-H), 5.32 (lH, t, J=8 Hz, 2"-~:, 4.93 (].H, d, J=8 H~, l"-H), 4.50 (lH, q, 7"-H), 1.31 (3H, d, J=5 Hz, 7"-CH3).
Anal Calcd for C51H4617 C 65-80~ H 4-55-Found: C 65.28, H 4.98.
Example 8. Preparation of 2"-0-benzoyletoposide (VIII, Rl=CH3, R2=H, R3=PhCO-, R4=H) A stirred solution of 4'-benzyloxycarbonyletoposide 2"-O-benzoate (54 mg, 0.07 mmol) in ethanol-acetone (2:1, 3 ml) was hydrogen~ted for 2 hours i.n the presenc~ of 10%
palladium on carbon (50 mg) at 1 atm. The catalyst was filtered off and the filtrate was concentrated to give the title compound (40 mg, 88% as colorless powder. Estimated purity 95% by HPLC.
MP ~89-292C.
IR ~max (KBr) cm 3450, 1760, 1730 UV ~max (MeOH) nm (~) 283 (4,470) H NMR (CDC13) ~ 7.4-7.8 (5H, m, COC6H5), 5 07 (1~, dd, J=7.7 & 9.2 Hz, 2"-H), 4.89 (lH, d, J=7 7 Hz, l"-H), 4.80 (lH, q, J=5.1 Hz, 7"-H), 4.23 (1~, dd, J=4.8 & 10.3 Hz, 6"-Heq), 3.98 (lH, dt, J=3.3 & 9.2 Hz, 3"-H), 3.64 (lH, t, J=10.3 Hz, 6"-Hax), 3.47 ~lH, t, J=9.2 .~z, 4"-H), 3.40 (lH, 8~Çi7 ''' dt, J=4.8 ~ 10 Hz, 5"-H), 2.74 (lH, d, J=3.3 Hz, 3"=OH), 1.41 (3H, d, J=5.1 Hz, 7"-CH3).
i~nal Calcd for C36H36Ql4 H20 Found: C 61.10, H 5.40.
Example 9. Preparation of 3"-0 Benzoyletoposide (VIII, R1=
CH3, R2=H, R3=H, R4=PhCO) According to the procedure of Example 8, 34 mg (C.04 mmol~ of 4'-0-benzyloxycarbonyletoposide 3"-0-benzoate was hydrogenated to give the title compound (27 mg, 95%) as colorless solid, which was purified on column chromatography to give colorless crystals. Estimated purity 95% by HPLC.
MP 188-191C.
IR `'max (KBr) cm 1 3400 (br), 1730.
UV ~max (MeOH) nm (E) ~23 (35,900), 280 (4,030).
H NMR (CDCl3) ~ 7.4-7.8 (5H, m, COPh), 5.38 ~lH, t, J=9.2 Hz, 3"-H), 4.79 (lH, d, J=7.7 Hz, l"-H), 4.72 (lH, q, J=5.1 Hz, 7"-H), 4.22 (lH, dd, J=5 & 10 Hz, 6"-Heq), 3.67 (lH, brt, J=8 Hz, 2"-H;, 3.6~ (lH, t, J=10 Hz, 6"-H~ax?~ 3.59 (lH, t, J=9.4 Hz, 4"-H), 3.47 (lH, m, 5"-H), 2..58 (lH, br, 2"-OH), 1.31 (3H, d, J=5.1 Hz, 7"-CH3).
Anal Calcd for C36H36O14 2H2O
Found: C 59.14, H 5.10.
Exam~le 10. Preparation of 2",3"-O-di-benzoyletoposide (VIII, R1=CH3, R2=H, R3=R4=PhCO-) A stirred solution of 4'-0-benzyloxycârbonyletoposide 2",3"-di-0-benzoate (130 mg, 0.14 mmol) in ethyl acetate-ethanol-acetone (4:4:1, 4.5 ml) was hydrogenated according to the procedu e of Example 8 to give the title compound (112 mg, ca. 100%) as colorless powder. E~'imateci ~0~ 67 purity 90% by HPLC.
MP >295C.
IR vmax (KBr) cm 1 1770, 1730.
UV ~max (MeOH) nm (E) 226 (sh, 24,200), 281, (3,470).
H NMR (CDCl3) ~ 7.3-8.0 (lOH, m, 2xCOC6H5), 5.67 (lH, t, J=9.5 Hz, 3"-H), 5.35 (lH, dd, J=8.1 & 9.7 Hz, 2"-H), 498 (lH, d, ~=8.1 Hz, l"-H), 4.73 (lH, q, J=4.8 Hz, 7"-H), 4.28 (lH, dd, J=4.4 & 10.3 Hz, 6"-Heq), 3.69 (lH, t, J=9.5 Hz, 4"-H), 3.67 (lH, t, J=10 Hz, 5"-Hax), 3.56 (lH, dt, J=9.5 &
lO H~, 5"-H), 1.33 (3H, d, J=4.8 Hz, 7"-CH3).
Anal Calcd for C43H400l5 H2 Found: C 62.99, H 4.99.
Example 11 The general procedure described in Example 1 is follo~!ed using 4'-CBZ teniposide instead of 4'-CBZ etoposide to provide 2",3"-di-0-acetyl teniposide.
Example 12 The general procedure described in Examples 2-4 is repeated using 4'-CBZ teniposide instead of 4'-CBZ etoposide to provide 2"-0-acetyl teniposide and 3"-0-teniposide.
Example 13 The gene-21 proces~lre s;essribed in ~x~.mF`e ~ i.s repeated usiilg ~'-C~Z teni?^s_dP insteas~ of ~ -C-Z etopos de ~o pro~.-ide 2",3"-di-0-formyl teniposide.
- 2~
ExamPle 14 The general procedure of Examples 7-10 is repeated using 4'-CBZ teniposide instead of 4'-CBZ etoposide to provide 2"-0-benzoyl teniposide, 3"-0-benzoyl teniposide and 2",3"-di-0-benzoyl teniposide.
Claims (13)
1. A compound having the formula wherein R2 is H and R1 is selected from the group consisting of (C1-10)alkyll (C2-10)alkenyl, (C5-6)cycloalkyl, 2-furyl,
2-thienyl, (C6-10)aryl, and (C7-14)aralkyl; or R1 and R2 are each (C1-10)alkyl; or R1, R2 and the carbon to which they are attached together represent (C5-6)cycloalkyl; one of R3 and R4 is H and the other is selected from the group consisting of (C1-5)alkanoyl and benzoyl; or R3 and R4 are the same and are selected from the group consisting of (C1-5) alkanoyl and benzoyl; R5 is H or a phosphate group.
2. A compound of Claim 1 wherein R2 is H and R1 is selected from the group consisting of methyl, 2-thienyl and phenyl.
2. A compound of Claim 1 wherein R2 is H and R1 is selected from the group consisting of methyl, 2-thienyl and phenyl.
3. A compound of Claim 2 wherein R1 to methyl.
4. A compound of Claim 1 wherein R3 and R4 are each (C1-5)alkanoyl
5. A compound of Claim 1 wherein one of R3 and R4 is H, and the other is (C1-5)alkanoyl or benzoyl.
6. A compound of Claim 4 wherein R2 is H and R1 is selected from the group consisting of methy and 2-thienyl.
7. A compound of Claim 6 wherein R1 is methyl, R3 and R4 are both formyl, and R5 is H.
8. A compound of Claim 6 wherein R1 is methyl, R3 and R4 are both acetyl, and R5 is H.
9. A compound of Claim 3 wherein R3 and R4 are both benzoyl, and R5 is H.
10. The compound of Claim 3 wherein R5 is H, one of R3 and R4 is H, and the other is (C2-5)alkanoyl.
11. The compound of Claim 3 wherein R5 is H, one of R3 and R4 is H, and the other is benzoyl.
12. A pharmaceutical composition comprising an antitumor effective amount of a compound of Claim 1 and a pharmaceutically acceptable carrier.
13. A method for inhibiting tumor growth in a mammalian host which comprises administering to said host an antitumor effective amount of a compound of Claim 1.
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| Application Number | Priority Date | Filing Date | Title |
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| US362,555 | 1989-06-07 | ||
| US07/362,555 US5036055A (en) | 1989-06-07 | 1989-06-07 | Acylated derivatives of etoposide |
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| US5191071A (en) * | 1987-08-21 | 1993-03-02 | Novo Nordisk A/S | Monoesters of glycosides and a process for enzymatic preparation thereof |
| US6610299B1 (en) | 1989-10-19 | 2003-08-26 | Aventis Pharma Deutschland Gmbh | Glycosyl-etoposide prodrugs, a process for preparation thereof and the use thereof in combination with functionalized tumor-specific enzyme conjugates |
| US7241595B2 (en) * | 1989-10-20 | 2007-07-10 | Sanofi-Aventis Pharma Deutschland Gmbh | Glycosyl-etoposide prodrugs, a process for preparation thereof and the use thereof in combination with functionalized tumor-specific enzyme conjugates |
| US6475486B1 (en) | 1990-10-18 | 2002-11-05 | Aventis Pharma Deutschland Gmbh | Glycosyl-etoposide prodrugs, a process for preparation thereof and the use thereof in combination with functionalized tumor-specific enzyme conjugates |
| US5034380A (en) * | 1989-11-20 | 1991-07-23 | Bristol-Myers Squibb Company | Alkoxymethylidene epipodophyllotoxin glucosides |
| CN1025918C (en) * | 1990-06-07 | 1994-09-14 | 国家医药管理局上海医药工业研究院 | Process for synthesis of antineoplastic medicine-etoposide |
| TW260671B (en) * | 1991-04-29 | 1995-10-21 | Bristol Myers Squibb Co | |
| IT1250692B (en) * | 1991-07-23 | 1995-04-21 | PROCEDURE FOR THE PREPARATION OF DEMETYLEPIPODOPHYLOTOXY-BETA-D-GLUCOSIDES. | |
| JP3061476B2 (en) * | 1992-04-24 | 2000-07-10 | 日本化薬株式会社 | Method for producing etoposide phosphate |
| US5459248A (en) * | 1993-11-04 | 1995-10-17 | Bristol-Myers Squibb Company | Process of preparing etoposide phosphate and etoposide |
| IL119749A (en) * | 1995-12-04 | 2001-03-19 | Nippon Kayaku Kk | Process for producing etoposide |
| AU728986B2 (en) * | 1997-06-02 | 2001-01-25 | Bristol-Myers Squibb Company | Process for preparing etoposide |
| CA2385528C (en) | 1999-10-01 | 2013-12-10 | Immunogen, Inc. | Compositions and methods for treating cancer using immunoconjugates and chemotherapeutic agents |
| US8240508B2 (en) | 2008-12-29 | 2012-08-14 | Gojo Industries, Inc. | Low cost radio frequency identification (RFID) dispensing systems |
| KR20150108391A (en) * | 2013-01-14 | 2015-09-25 | 어드바노믹스 코포레이션 | Gem-difluorinated c-glycoside compounds as anti-cancer agents |
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| NL6613143A (en) * | 1965-09-21 | 1967-03-22 | ||
| CH469000A (en) * | 1965-12-14 | 1969-02-28 | Sandoz Ag | Process for the production of new glucosides |
| FR1518706A (en) * | 1965-12-14 | 1968-03-29 | Sandoz Sa | Glucosides derived from podophyllotoxin related compounds and their preparation |
| CH514578A (en) * | 1968-02-27 | 1971-10-31 | Sandoz Ag | Process for the production of glucosides |
| JPS58219196A (en) * | 1982-06-15 | 1983-12-20 | Nippon Kayaku Co Ltd | Production of 4'-demethyl-epipodophyllotoxin-beta-d- ethylideneglucoside |
| JPS58225096A (en) * | 1982-06-25 | 1983-12-27 | Nippon Kayaku Co Ltd | Novel preparation of 4'-demethyl-epipodophyllotoxin-beta- d-ethylideneglucoside |
| DE3374317D1 (en) * | 1982-11-26 | 1987-12-10 | Nippon Kayaku Kk | Process for producing 4'-demethyl-epipodophyllotoxin-beta-d-ethylidene-glucoside and acyl-derivative thereof |
| US4567253A (en) * | 1984-02-03 | 1986-01-28 | Tony Durst | 2-Substituted derivatives of podophyllotoxin and etoposide |
| JPS60246393A (en) * | 1984-05-22 | 1985-12-06 | Nippon Kayaku Co Ltd | Novel preparation of etoposide |
| US4609644A (en) * | 1984-06-15 | 1986-09-02 | St. Jude Children's Research Hospital | Epipodophyllotoxinquinone glucoside derivatives, method of production and use |
| JPS61134396A (en) * | 1984-12-06 | 1986-06-21 | Nippon Kayaku Co Ltd | Novel production of etoposide |
| JPS61227590A (en) * | 1985-04-02 | 1986-10-09 | Microbial Chem Res Found | Novel 4'-epopodophyllotoxin derivative |
| IL77334A (en) * | 1985-12-16 | 1991-04-15 | Univ Bar Ilan | Synthesis of 9-epipodophyllotoxin glucoside derivatives and some novel intermediates therefor |
| JPS63150293A (en) * | 1986-12-12 | 1988-06-22 | Nippon Kayaku Co Ltd | Water-soluble etoposide derivative |
| US4916217A (en) * | 1987-01-08 | 1990-04-10 | Bristol-Myers Company | Phosphorus containing derivatives of epipodophyllotoxin |
| JPS63192793A (en) * | 1987-02-06 | 1988-08-10 | Nippon Kayaku Co Ltd | Novel ester of 4'-demethyl-epipodophyllotoxin derivative |
| US4853467A (en) * | 1987-05-19 | 1989-08-01 | Bristol-Myers Company | Nitrogen containing derivatives of epipodophyllotoxin glucosides |
| US4874851A (en) * | 1987-07-01 | 1989-10-17 | Bristol-Meyers Company | 3',4'-dinitrogen substituted epipodophyllotoxin glucoside derivatives |
| US4904768A (en) * | 1987-08-04 | 1990-02-27 | Bristol-Myers Company | Epipodophyllotoxin glucoside 4'-phosphate derivatives |
| US4868291A (en) * | 1987-08-20 | 1989-09-19 | Bristol-Myers Company | 4'-deshydroxyepipodophyllotoxin glucosides: synthesis and use |
| US4888419A (en) * | 1987-08-31 | 1989-12-19 | Bristol-Myers Company | 3'-demethoxyepipodophyllotoxin glucoside derivatives |
| ZA886810B (en) * | 1987-12-18 | 1989-08-30 | Bristol Myers Co | Epipodophyllotoxin glucoside 4'-acyl derivatives |
| US4912204A (en) * | 1988-09-06 | 1990-03-27 | Bristol-Myers Company | Fluoro-substituted epipodophyllotoxin glucosides |
| US5066645A (en) * | 1989-09-01 | 1991-11-19 | Bristol-Myers Company | Epipodophyllotoxin altroside derivatives |
-
1989
- 1989-06-07 US US07/362,555 patent/US5036055A/en not_active Expired - Fee Related
-
1990
- 1990-06-01 NZ NZ233900A patent/NZ233900A/en unknown
- 1990-06-01 CA CA002018067A patent/CA2018067A1/en not_active Abandoned
- 1990-06-04 IL IL94606A patent/IL94606A0/en unknown
- 1990-06-04 CN CN90104190A patent/CN1024197C/en not_active Expired - Fee Related
- 1990-06-04 FI FI902767A patent/FI902767A0/en not_active Application Discontinuation
- 1990-06-05 KR KR1019900008275A patent/KR910000778A/en not_active Application Discontinuation
- 1990-06-06 PT PT94281A patent/PT94281A/en not_active Application Discontinuation
- 1990-06-06 AU AU56828/90A patent/AU637104B2/en not_active Ceased
- 1990-06-06 YU YU01106/90A patent/YU110690A/en unknown
- 1990-06-06 EP EP19900110729 patent/EP0401800A3/en not_active Ceased
- 1990-06-06 ZA ZA904366A patent/ZA904366B/en unknown
- 1990-06-06 NO NO902495A patent/NO173239C/en unknown
- 1990-06-06 DD DD90341368A patent/DD294947A5/en not_active IP Right Cessation
- 1990-06-07 JP JP2147535A patent/JPH0324097A/en active Pending
- 1990-06-07 HU HU903517A patent/HU206221B/en not_active IP Right Cessation
- 1990-06-07 CZ CS902838A patent/CZ283890A3/en unknown
Also Published As
| Publication number | Publication date |
|---|---|
| CN1047869A (en) | 1990-12-19 |
| AU637104B2 (en) | 1993-05-20 |
| US5036055A (en) | 1991-07-30 |
| EP0401800A2 (en) | 1990-12-12 |
| EP0401800A3 (en) | 1991-04-10 |
| NO902495L (en) | 1990-12-10 |
| CZ283890A3 (en) | 1995-02-15 |
| NO902495D0 (en) | 1990-06-06 |
| PT94281A (en) | 1991-02-08 |
| KR910000778A (en) | 1991-01-30 |
| NO173239C (en) | 1993-11-17 |
| YU110690A (en) | 1992-05-28 |
| ZA904366B (en) | 1991-05-29 |
| NO173239B (en) | 1993-08-09 |
| FI902767A0 (en) | 1990-06-04 |
| IL94606A0 (en) | 1991-04-15 |
| AU5682890A (en) | 1990-12-13 |
| CN1024197C (en) | 1994-04-13 |
| HU206221B (en) | 1992-09-28 |
| HU903517D0 (en) | 1990-10-28 |
| NZ233900A (en) | 1992-02-25 |
| DD294947A5 (en) | 1991-10-17 |
| HUT54174A (en) | 1991-01-28 |
| JPH0324097A (en) | 1991-02-01 |
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Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| FZDE | Discontinued |